| Acute lung injury/acute respiratory distress syndrome(ALI/ARDS) is characterized by a pulmonary inflammation response that a number of neutrophils(PMN) infiltrated into the lungs. PMN and other cell accumulation in the lungs, increased generation of various enzyme and reactive oxygen intermediates(ROI), and the release of inflammatory mediators mainly contribute to the pulmonary damage. PMN has been identified to play a key role in ALI, however, the mechanism how the leading PMN was activated in lung is unknown at present.One family of intracellular messengers, found in all mammalian cells inclunding PMN, is the mitogen activated protein kinase(MAPK),a serine-threonine-directed kinase activated by phosphorylation. MAPK activation has been implicated in many physiological responses process, which lead the stimuli from membrane to nucleus.There are 3 members of the MAPK family:extracellular signal-regulated kinase(ERK), P38MAPK, and c-Jun N-terminal kinase(JNK). Recent studies show that the MAPK in PMN, particularly ERK, could be activated by diverse extracellular stimuli, including mitogens, growth factors, oxidative stress, and cytokines. After activation, ERK pathway will involve in modulating the activation of transcriptional factors such as cAMP response element-binding protein (CREB), nuclear factor - IL - 6 (NF-IL-6), activating transcription factor-2 and AP-1, then inluence PMN functions and apoptosis. This suggest that MAPK activation in PMN playsan important role during the course of the disease. Therefore, we investigated the relation of the delay for PMN apoptosis in ALI bronchoalveolar lavage fluid(HALF) with MARK activation and inflammation mediator production in lungs in order to elucidate the role for Raf-1→MEK-1→MAPK signal transduction system in ALI.In the present experiment, the murine model of ALI was estabalished by instillation of little acid into lungs and subsequent lipopolysaccharide(LPS) 24hrs after acid instillation. In contrast, the control group were instilled by aseptic injection water. After above manipulations, BAL was perfomed at different times in experimental and control groups. And the BALF was spun. Total cell counts and cell differential counts were performed. Wet to dry weight ratio was measured. After normal peripheral blood PMNs separated from the control were incubated differently with the ALI and control BALF supernatant 16 to 24h, the apoptosis of PMN was analyzed by Annexin V binding using flow cytometry. ERK and phosphorylated ERK(P-ERK) in each group PMN were determined by Western blot. Simultaneously, lung tissue was fixed for histologic examination. Further more, the activation of Raf-1→MEK-1 →MAPK signal transduction system and iNOS, TNF-α expression were determined by immunohistochemistry method.Results:(1)Respiratory rate in the experimental rats obviously increased after LPS instillation, but the sham control have no abnormal appearance. The PMN count in ALI BALF and wet to dry weight ratio of lung were significantly increased compared with control. The histologic appearance of the lungs from ALI rats includes thickening of thealveolar septum, increased interstitial edema, abroadly dispersing inflammation, and the infiltration of a large number PMNs and red cells into alveolar spaces as well as alveolar septum. In contrast, none of the changes was seen in control group. (2)The apoptosis of normal peripheral blood PMN incubated in control BALF supernatant was low?0%) over the first 8h, but a exponential increase in apoptosis occurred between 8 and 24hrs , and the proportion of apoptosis was the highest at 24h. In contrast, when normal PMN were incubated in ALI BALF supernatant, the proportion of apoptotic PMN was significantly lower compared with incubation in control BALF(p<0.01), although apoptosis occurred to a degree after 16h. In addition, the percentage of apoptotic PMN in ALI BALF was remarkably lower at 16 and 24h, compared with normal PMN incubated in control BALF over 16 to 24hrs(p<0. 01), and no differences were noted between...
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